PROJECT SUMMARY/ABSTRACT This project will study how glucocorticoids (GCs) interact with glucocorticoid receptors (GRs) in sensory neurons and glia to effect the development and maintenance of pain after nerve injury. The majority of studies examining nerve-injury induced pain do not examine where GC/GR interactions are important for pain development and or how they contribute to the changes that arise after injury (e.g. reactive gliosis). Thus, this project is relevant to the NIH mission because it examines the fundamental importance of GRs in various nervous system cell types and will elucidate where GR expression is required for the development of neuronal hyperexcitability and pain after injury. Even though GR expression in the spinal cord increases after injury, whether GR activity is necessary for downstream responses has not been demonstrated. Thus we will use Cre-Lox systems to evaluate GR loss in sensory neurons, microglia and astrocytes with and without elevated corticosterone levels to determine where GC/GR interactions are influencing pain and neuronal excitability. Extensive sensory behavior testing will determine if loss of GR in each cell type effects reflexive and spontaneous pain development and maintenance. Three-dimensional tissue clearing techniques will provide an overarching view of neuronal excitability changes using cFOS in the spinal cord and brain, as well as facilitate the visualization of changes in sensory process innervation in the periphery and in the CNS. Changes in sensory neuron excitability will be directly measured using patch-clamp recording. Glial activation, gene expression, and morphology will be extensively examined in each condition. Together, these data will provide fundamental insight into glucocorticoid receptor biology in sensory neurons, microglia, and astrocytes as well as discern the importance of GR actions in neurons versus glia in an injury environment.